A competitive protein binding assaywas used for the determination of adenosine 3′,5′-monophosphate (cyclic AMP) in human amniotic fluid. A cyclic AMP specific binding protein prepared from rabbit skeletal muscle was utilized and the method was applicable for accurate measurements of this nucleotide in the 0.1–1.0 pmole range. The levels of cyclic AMP in amniotic fluid samples from various stages of normal pregnancies were measured and the values ranged from 2.6 pmoles per ml in the fluid at 4 weeks gestation to an average of 19.2 pmoles per ml in the 40th week of pregnancy. The lowest cyclic AMP levels were found in amniotic fluids in the 4–14 weeks gestational period (2.8 ± 0.6 (mean ± sd) pmoles per ml). Thereafter, the fluid cyclic AMP concentration showed a steady increase with the advance of pregnancy, reaching a peak value in the fluids at term. The mean level during the 16–24 weeks gestation period was 4.9 ± 0.8 (mean ± sd) pmoles per ml and a maximum level of 17.6 ± 3.6 (mean ± sd) was found in the fluids at 34–40 weeks gestation. In addition, attempts were made to associate cyclic AMP level in the fluids from term pregnancy to various postpartum status of the respective infants. A positive correlation was established between the fluid cyclic AMP level and the corresponding birth weight of the newborn. The combined results indicated that changes in the amniotic fluid cyclic AMP levels may be related to certain aspects in fetal development and that the empirical relationship between the fluid cyclic AMP concentration and fetal growth may have useful clinical applications.